Respirable crystalline silica mineral particles inhaled through workplace exposure are an important world-wide cause of chronic lung disease and pulmonary fibrosis. The goal of this research is to delineate the cytokine-driven mechanisms of lung scarring caused by the xenobiotic agent silica. The proposal uses genetically modified cytokine deficient strains of mice, and normal mice with cytokines blocked or augmented, to determine the mechanisms of silica-induced fibrosis in a whole-body inhalation model of silicosis. The animal experiments will be extended by a simplified in vitro paradigm of silicosis, a cell culture system utilizing silicotic pulmonary macrophages and target lung fibroblast cell lines isolated from wild-type mice and mutant strains with cytokine receptor deletions. Interferon-gamma (IFN- gamma) from lymphocytes is the predominant cytokine that drives macrophage activation. IFN-gamma is abundant in silicotic lesions in mouse lungs and its removal in IFN-gamma gene-deleted (knockout) mice results in less silicosis and pulmonary fibrosis after silica inhalation. Interleukin-12 (IL-12) is the major macrophage-derived cytokine that drives lymphocytes to IFN-gamma production. We hypothesize that macrophages with direct silica contact are stimulated to produce IL-12, that initial IFN-gamma production activates a recruited, expanded macrophage population to produce cytokines that promote fibrosis (e.g. PDGF, TNF-alpha, IGF-1, TGF-beta), and re-iteratively induce additional IL-12 and IFN-gamma production. We hypothesize that IFN-gamma also may directly modify the effects of these cytokines on lung fibroblasts to control the extent of fibrosis. Our Specific Aims are (1) To determine how interferon-gamma (IFN-gamma) increases the extent of pulmonary fibrosis caused by silica, (2) To determine if interleukin-12 (IL-12) is a pivotal control point through which silicotic macrophages stimulate lymphocyte IFN- gamma production, and (3) To define the downstream cytokine signaling cascade through which IFN-gamma may modify the fibrogenic effects of silicotic macrophages on lung fibroblasts. We will achieve these aims using a cross-confirming strategy in which we will (a) remove IFN-gamma using IFN-gamma knockout mice, neutralizing antibody, or IFN-gamma receptor deficient target cells, (b) augment IFN-gamma using recombinant exogenous IFN- gamma, or (c) replace IFN-gamma in IFN-gamma knockout mice using, rmIFN-gamma. A similar approach will study IL-12. Understanding the mechanisms through which this toxic agent causes pulmonary fibrosis will be a significant advance, and may offer intervention opportunities for silicosis. And other interstitial lung diseases where the cause is not known and treatment is not effective.